protein synthesis notes ch 17 central dogma dna rna protein

Protein Synthesis Notes

Ch 17

Central Dogma

• DNA RNA Protein

Prokaryotic vs Eukaryotic Transcription

Prokaryotic Transcription-takes place in cytoplasm -circular DNA w/no histones-no introns / no mRNA processing

Eukaryotic Transcription -takes place in nucleus -DNA has histones and linear-introns and mRNA processing

Prokaryotic Transcription

1. Initiation – RNA polymerase binds to promoter

2. Elongation – RNA polymerase reads template 3’-5’ and builds mRNA strand 5’-3’

3. Termination – mRNA is released …forms hairpin loop.

STEPS

Eukaryotic Transcription1. Initiation – Transcription factors

adhere to the TATA box in the promoter signaling RNA Polymerase II to attach. Additional transcription factors attach and the transcription initiation complex is formed. Enhancers/silencers

2. Elongation – RNA Polymerase II unzips the DNA and pairs the template withcomplementary mRNA nucleotides.

3. Termination – RNA Polymerase II reaches the polyadenylation (AAUAAA)

sequence and releases the pre-mRNA.

*Gene expression is most often regulated at transcription

transcription

Eukaryotic TranscriptionHHMI transcription initiation complex

Transcription

mRNA processing

mRNA Processing (takes place in the nucleus)1. Add 5’ cap (guanine with 3 phosphates) 2. Add Poly A tail (repeated Adenines)3. Introns (non coding regions) cut out4. Exons (coding regions) fused together

snRNPs (small nuclear ribonucleoproteins)

• snRNPs- small nuclear RNA– RNA and proteins

• Spliceosome - several snRNPs that carry out RNA splicing

Spliceosome Animation

RibozymeRibozyme – RNA that functions as an enzyme-splicing without proteins!

How: 1. Single stranded and forms 3-D structure2. Bases in RNA contain functional groups that may catalyze rxns. 3. Forms hydrogen bonds

Sidney Altman Thomas CechYale U of Colorado

Harvard Ribozymes

Thomas-Cech Ribozymes

Animations• http://www-class.unl.edu/biochem/gp2/m_biology/animation/gene/gene_a2.html• http://www.stolaf.edu/people/giannini/flashanimat/molgenetics/transcription.swf• http://www.concord.org/~btinker/workbench_web/models/eukTranscription.swf• http://vcell.ndsu.edu/animations/transcription/index.htm• http://www.hhmi.org/biointeractive/dna/animations.html

2007-2008

Translationfrom

nucleic acid languageto

amino acid language

Translation • Codons– 3 nucleotides

decoded into the sequence of amino acids

2007-2008

Translation in Prokaryotes

Bacterial chromosome

mRNA

Cell wall

Cellmembrane

Transcription

Translation

proteinPsssst…no nucleus!

• Transcription & translation are simultaneous in bacteria – DNA is in

cytoplasm– no mRNA

editing – ribosomes

read mRNA as it is being transcribed

Translation in Prokaryotes

Translation: prokaryotes vs. eukaryotes• Differences between prokaryotes &

eukaryotes– time & physical separation between processes• takes eukaryote ~1 hour

from DNA to protein– RNA processing

2007-2008

Translation in Eukaryotes

mRNA

From gene to protein

DNAtranscription

nucleus cytoplasm

mRNA leaves nucleus through nuclear pores

proteins synthesized by ribosomes using instructions on mRNA

aa

aa

aaaa

aa

aaaa

aa

ribosome

proteintranslation

How does mRNA code for proteins?TACGCACATTTACGTACGCGG

DNA

AUGCGUGUAAAUGCAUGCGCC

mRNA

Met Arg Val Asn Ala Cys

Alaprotein

?

4

4

20

ATCG

AUCG

AUGCGUGUAAAUGCAUGCGCC

mRNA

mRNA codes for proteins in triplets

TACGCACATTTACGTACGCGG

DNA

AUGCGUGUAAAUGCAUGCGCCmRNA

Met Arg Val Asn Ala

Cys Ala

protein

?

codon

The code• Code for ALL life!

– strongest support for a common origin for all life

• Code is redundant– several codons for each

amino acid– 3rd base “wobble”

Start codon AUG methionine

Stop codons UGA, UAA, UAG

Why is thewobble good?

2007-2008

Translationfrom

nucleic acid languageto

amino acid language

Translation • Codons– blocks of 3

nucleotides decoded into the sequence of amino acids

2007-2008

Translation in Prokaryotes

Bacterial chromosome

mRNA

Cell wall

Cellmembrane

Transcription

Translation

proteinPsssst…no nucleus!

• Transcription & translation are simultaneous in bacteria – DNA is in

cytoplasm– no mRNA

editing – ribosomes

read mRNA as it is being transcribed

Translation in Prokaryotes

Translation: prokaryotes vs. eukaryotes• Differences between prokaryotes &

eukaryotes– time & physical separation between processes• takes eukaryote ~1 hour

from DNA to protein– RNA processing

2007-2008

Translation in Eukaryotes

mRNA

From gene to protein

DNAtranscription

nucleus cytoplasm

mRNA leaves nucleus through nuclear pores

proteins synthesized by ribosomes using instructions on mRNA

aa

aa

aaaa

aa

aaaa

aa

ribosome

proteintranslation

How does mRNA code for proteins?TACGCACATTTACGTACGCGG

DNA

AUGCGUGUAAAUGCAUGCGCC

mRNA

Met Arg Val Asn Ala Cys

Alaprotein

?

4

4

20

ATCG

AUCG

AUGCGUGUAAAUGCAUGCGCC

mRNA

mRNA codes for proteins in triplets

TACGCACATTTACGTACGCGG

DNA

AUGCGUGUAAAUGCAUGCGCCmRNA

Met Arg Val Asn Ala

Cys Ala

protein

?

codon

The code• Code for ALL life!

– strongest support for a common origin for all life

• Code is redundant– several codons for each

amino acid– 3rd base “wobble”

Start codon AUG methionine

Stop codons UGA, UAA, UAG

Why is thewobble good?

How are the codons matched to amino acids?

TACGCACATTTACGTACGCGGDNA

AUGCGUGUAAAUGCAUGCGCCmRNA

aminoacid

tRNA

anti-codon

codon

5 3

3 5

3 5

UAC

MetGCA

ArgCAU

Val

mRNA

From gene to protein

DNAtranscription

nucleuscytoplasm

aa

aa

aaaa

aa

aaaa

aa

ribosome

proteintranslation

aa

Transfer RNA structure• “Clover leaf” structure– anticodon on “clover leaf” end– amino acid attached on 3 end

tRNA – “Wobble”

Inosine can pair with C, A, U allowing for less tRNA’s, More relaxed at the 3rd base position.

Loading tRNA • Aminoacyl tRNA synthetase – enzyme which bonds amino acid to tRNA– bond requires energy

• ATP AMP– energy stored in tRNA-amino acid bond

• unstable• so it can release amino acid at ribosome easily

activatingenzyme

anticodontRNATrp binds to UGG condon of mRNA

Trp Trp Trp

mRNAA C CU G G

C=OOH

OHH2OO

tRNATrp

tryptophan attached to tRNATrp

C=O

O

C=O

Ribosomes • Facilitate coupling of

tRNA anticodon to mRNA codon

• Structure– ribosomal RNA (rRNA) & proteins– 2 subunits• large• Small

E P AAssembled in the nucleus of Eukaryotes

Ribosomes

Met

5'

3'

UUA C

A G

APE

• A site (aminoacyl-tRNA site) – holds tRNA carrying next amino acid to be

added to chain • P site (peptidyl-tRNA site) – holds tRNA carrying growing polypeptide chain

• E site (exit site)– empty tRNA

leaves ribosome from exit site

Translation Animation McGraw HillUconn Translation Animation

Building a polypeptide

• Initiation– brings together mRNA, ribosome

subunits, initiator tRNA

• Elongation– adding amino acids based on codon

sequence

• Termination– end codon 123

Leu

Leu Leu Leu

tRNA

Met MetMet Met

PE AmRNA

5' 5' 5' 5'3' 3' 3' 3'

U UA A AACC

CA U UG G

GUU

A AAAC

CC

A U UG GGU

UA A A

ACC

CA U UG G

GU UA A ACCA U UG G

G A C

Val Ser

Ala Trp

releasefactor

AA A

C CU UG G 3'

Protein targeting • Signal peptide– address label

Destinations: secretion nucleus mitochondria chloroplasts cell membrane cytoplasm etc…start of a secretory pathway

Can you tell the story?

DNA

pre-mRNA

ribosome

tRNA

aminoacids

polypeptide

mature mRNA

5' cap

polyA tail

large ribosomal subunit

small ribosomal subunit

aminoacyl tRNAsynthetase

E P A

5'

3'

RNA polymerase

exon introntRNA